Various publications, including patents, published applications, technical articles and scholarly articles are cited throughout the specification. Each of these cited publications is incorporated by reference herein, in its entirety and for all purposes.
Breast cancer (BCa) is the most common cancer diagnosis in women and the second-leading cause of cancer-related death among women (Ries L A G, et al. (eds). SEER Cancer Statistics Review, 1975-2003, National Cancer Institute, Bethesda, Md.). Major advances in breast cancer treatment over the last 20 years have led to significant improvement in the rate of disease-free survival (DFS). For example, therapies utilizing antibodies reactive against tumor-related antigens have been used to block specific cellular processes in order to slow disease progress or prevent disease recurrence. Despite the recent advances in breast cancer treatment, a significant number of patients will ultimately die from recurrent disease. Thus, there is a need for treatments that prevent or slow or prohibit the development of recurrent disease.
Vaccines are an attractive model for such treatments and preventions due to their ease of administration, and because of their high rate of success observed for infectious diseases. The basic concept of constructing a cancer vaccine is straightforward in theory. The development of effective cancer vaccines for solid tumors in practice, however, has met with limited success. For example, one group attempting to administer a peptide vaccine directed against metastatic melanoma observed an objective response rate of only 2.6% (Rosenberg S A et al. (2004) Nat. Med. 10:909-15).
There are many potential explanations for this low success rate (Campoli M et al. (2005) Cancer Treat. Res. 123:61-88). For example, even if an antigen is specifically associated with a particular type of tumor cell, the tumor cells can express only low levels of the antigen, or it can be located in a cryptic site or otherwise shielded from immune detection. In addition, tumors often change their antigenic profile by shedding antigens as they develop. Also contributing to the low success rate is the fact that tumor cells can express very low levels of MHC proteins and other co-stimulatory proteins necessary to generate an immune response.
Additional problems facing attempts at vaccination against tumors arise in patients with advanced-stage cancers. Such patients tend to have larger primary and metastatic tumors, and the cells on the interior of the tumor can not be accessible due to poor blood flow. This is consistent with the observation that vaccine strategies have tended to be more successful for the treatment of hematologic malignancies (Radford K J et al. (2005) Pathology 37:534-50; and, Molldrem J J (2006) Biol. Bone Marrow Transplant. 12:13-8). In addition, as tumors become metastatic, they can develop the ability to release immunosuppressive factors into their microenvironment (Campoli, 2005; and, Kortylewski M et al. (2005) Nature Med. 11:1314-21). Metastatic tumors have also been associated with a decrease in the number of peripheral blood lymphocytes, and dendritic cell dysfunction (Gillanders W E et al. (2006) Breast Diseases: A Year Book and Quarterly 17:26-8).
While some or all of these factors can contribute to the difficulty in developing an effective preventative or therapeutic vaccine, the major underlying challenge is that most tumor antigens are self antigens or have a high degree of homology with self antigens, and are thus expected to be subject to stringent immune tolerance. Thus, it is clear that many peptide-based cancer vaccines, with or without immune-stimulating adjuncts, can be doomed to only limited success in clinical practice due to low immunogenicity and lack of specificity.
Prototype breast cancer vaccines based on single antigens have been moderately successful in inducing a measurable immune response in animal experiments and in clinical tests with breast cancer patients. The observed immune response, however, has not translated into a clinically-significant protective immunity against resurgence of disease put in remission by standard surgery and chemotherapy. Thus, novel vaccine approaches are needed to further improve recurrence rates and overall survival among BCa patients.
Preferred vaccine epitopes are those that are expressed exclusively, or at least at increased levels by a neoplasm. HER2/neu is a proto-oncogene expressed in many epithelial malignancies (Slamon D J et al. (1989) Science 244:707-12). Gene amplification and overexpression of the HER2/neu protein is found in 20-25% of BCa, and its excess presence is an indicator of poor prognosis (Pritchard K I et al. (2006) N. Engl. J. Med. 354:2103-11). HER2/neu has been studied fairly extensively, and several immunogenic peptides have been identified from this protein. One such peptide is termed E75, and corresponds to amino acids 369-377 of HER2/neu (SEQ ID NO:1) (U.S. Pat. No. 6,514,942).
Attempts have been made to utilize E75 as an anti-cancer vaccine, for example, as a single peptide vaccine combined with different immunoadjuvants (Zaks T Z et al. (1998) Cancer Res. 58:4902-8; Knutson K L et al. (2002) Clin. Cancer Res. 8:1014-8; and, Murray J L et al. (2002) Clin. Cancer Res. 8:3407-18); loaded on to autologous dendritic cells and reinfused (Brossart P et al. (2000) Blood 96:3102-8; and, Kono K et al. (2002) Clin. Cancer Res. 8:3394-3400); or embedded in longer peptides capable of binding HLA class II molecules in order to recruit CD4 helper T-cells (Disis M L et al. (1999) Clin. Cancer Res. 5:1289-97; and, Disis M L et al. (2002) J. Clin. Oncol. 20:2624-32). Each approach has stimulated an E75-specific cytotoxic T cell-mediated immune response, but has not demonstrated a clinically significant therapeutic or protective immunity in women with advanced stage breast cancer.
HER2/neu is a member of the epidermal growth factor receptor family and encodes a 185-kd tyrosine kinase receptor involved in regulating cell growth and proliferation. (Popescu N C, King C R, Kraus M H. Localization of the human erbB-2 gene on normal and rearranged chromosome 17 to bands q12-21.32. Genomics 1989; 4:362-366; Yarden Y, Sliwkowski M X. Untangling the ErbB signaling network. Nat Rev Mol Cell Bio 2001; 2:127-137.) Over-expression and/or amplification of HER2/neu is found in 25-30% of invasive breast cancers (BCa) and is associated with more aggressive tumors and a poorer clinical outcome. (Slamon D J, Clark G M, Wong S G, et al. Human breast cancer correlation of relapse and survival with amplification of the HER-2/neu oncogene. Science 1987; 235:177-182; Slamon D J, Godolphin W, Jones L A, et al. Studies of the HER-2/neu proto-oncogene in human breast and ovarian cancer. Science 1989; 244:707-12; Toikkanen S, Helin H, Isola J, Joensuu H. Prognostic significance of HER-2 oncoprotein expression in breast cancer: A 30-year follow-up. J Clin Oncol 1992; 10:1044-1048.)
Determining HER2/neu status is performed predominately via two tests, immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH). IHC detects over-expression of HER2/neu protein and is reported on a semi-quantitative scale of 0 to 3+ (0=negative, 1+=low expression, 2+=intermediate, and 3+=over-expression). FISH on the other hand detects amplification (excess copies) of the HER2/neu gene and is expressed as a ratio of HER2/neu gene copies to chromosome 17 gene copies and interpreted as “over-expression” if FISH is ≧2.0 copies. (Hicks D G, Tubbs R R. Assessment of the HER2 status in breast cancer by fluorescence in situ hybridization: a technical review with interpretive guidelines. Hum Pathol 2005; 36:250-261.) Concurrence rate of IHC and FISH is approximately 90%. (Jacobs T W, Gown A M, Yaziji H, et al. Specificity of HercepTest in determining HER-2/neu status of breast cancers using the United States Food and Drug Administration-approved scoring system. J Clin Oncol 1999; 17:1533-1541.) FISH is considered the gold standard, as retrospective analysis reveals it is a better predictor of trastuzumab (Tz) response; it is more objective and reproducible. (Press M F, Slamon D J, Flom K J, et al. Evaluation of HER-2/neu Gene Amplification and Overexpression: Comparison of Frequently Used Assay Methods in a Molecularly Characterized Cohort of Breast Cancer Specimens. J Clin Oncol 2002; 14:3095-3105; Bartlett J, Mallon E, Cooke T. The clinical evaluation of HER-2 status: which test to use? J Pathol 2003; 199:411-417; Wolff A C, Hammond M E H, Schwartz J N, et al. American Society of Clinical Oncology/College of American Pathologists guideline recommendations for human epidermal growth factor receptor 2 testing in breast cancer. J Clin Oncol 2007; 25:118-145.)
Identification and quantification of HER2/neu as a proto-oncogene has led to humoral or antibody-based passive immunotherapy, to include the use of Tz (Herceptin®). Tz is a recombinant, humanized monoclonal antibody that binds the extracellular juxtamembrane domain of HER2/neu protein. (Plosker G L, Keam S J. Trastuzumab: A review of its use in the management of HER2-positive metastatic and early-stage breast cancer. Drugs 2006; 66:449-475.) Tz is indicated for HER2/neu over-expressing (IHC 3+ or FISH ≧2.0) node-positive (NP) and metastatic BCa patients, (Vogel C L, Cobleigh M A, Tripathy D, et al. Efficacy and safety of trastuzumab as a single agent in first-line treatment of HER2-overexpressing metastatic breast cancer. J Clin Oncol 2002; 20:719-726; Piccart-Gebhart M J, Procter M, Leyland-Jones B, et al. Trastuzumab after adjuvant chemotherapy in HER2-positive breast cancer. N Engl J Med 2005; 353:1659-1672) and shows very limited activity in patients with low to intermediate HER2/neu expression. (Herceptin (Trastuzumab) prescription product insert. South San Francisco, Calif.: Genentech Inc; revised September 2000.)
Another form of immunotherapy being pursued is vaccination and active immunotherapy targeting a cellular immune response to epitopes on tumor associated antigens (TAA) such as HER2/neu. HER2/neu is a source of several immunogenic peptides that can stimulate the immune system to recognize and kill HER2/neu-expressing cancer cells. (Fisk B, Blevins T L, Wharton J T, et al. Identification of immunodominant peptide of the HER2/neu proto-oncogene recognized by ovarian tumor-specific CTL lines. J Exp Med 1995; 181:2109-2117.)
E75 (KIFGSLAFL, HER2/neu, 369-377) is a peptide sequence in the HER2/neu proto-oncogene family and is in use in clinical trials as an anti-cancer vaccine to stimulate cytotoxic T lymphocytes (CTL) to destroy cancer cells. (Zaks, T. et. al. Immunization with a peptide epitope (369-377) from HER-2/neu leads to peptide specific cytotoxic T lymphocytes that fail to recognize HER-2/neu+ tumors. Cancer Research. 58 (21): 4902-8. 1998; Knutson K L, Schiffman K, Cheever M A, et al: Immunization of cancer patients with HER-2/neu, HLA-A2 peptide, p369-377, results in short-lived peptide-specific immunity. Clin Cancer Res 8:1014-1018, 2002; Murray J L, Gillogly M E, Przepiorka D, et al: Toxicity, immunogenicity, and induction of E75-specific tumorlytic CTLs by HER-2 peptide E74 (369-377) combined with granulocyte macrophage colony-stimulating factor in HLA-A2+ patients with metastatic breast and ovarian cancer. Clin Cancer Res 8:3407-3418, 2002; Avigan D, Vasir B, Gong J, et al. Fusion cell vaccination of patients with metastatic breast and renal cell cancer induces immunological responses. Clin Cancer Res 2004: 10:4699-4708; Disis M L, Gooley T A, Rinn K, et al. Generation of T-cell immunity to the HER2/neu protein after active immunization with HER2/neu peptide-based vaccines. J Clin Oncol 2002; 20:2624-32; Disis M L, Grabstein K H, Sleath P R, et al. Generation of immunity to the HER-2/neu oncogenic protein in patients with breast and ovarian cancer using a peptide-based vaccine. Clin Cancer Res 5:1289-1297, 1999.
Targeted passive immunotherapy based on the HER2/neu proto-oncogene has primarily revolved around the use of Tz (Herceptin®). Tz is a recombinant, humanized monoclonal antibody that binds the extracellular juxtamembrane domain of the HER2/neu protein. Tz is approved by regulatory authorities and indicated for treatment of HER2/neu over-expressing (IHC 3+ or FISH >2.0) tumors in metastatic breast cancer patients and in the adjuvant setting for node-positive breast cancer patients. Tz has undergone multiple clinical trials and is now routinely used in the treatment of metastatic patients and in the adjuvant treatment of high risk breast cancer patients with overexpression of HER2/neu. Tz, however, shows limited activity in patients with low to intermediate HER2/neu expression. Therefore, based on the previous results seen with Tz, immunogenic peptide vaccines targeting HER2/neu would not be expected to be effective in cancer patients with low and intermediate levels of HER2/neu tumor expression.
Thus, there is a need in the art to exploit the immunoprotective and therapeutic potential of E75 to produce vaccines that offer breast cancer patients in clinical remission reliable protection against recurrence of the disease.